Electrophysiologic studies indicate that excitatory opioid receptors on mouse sensory dorsal-root ganglion (DRG) neurons in culture are positively coupled via Gs to cyclic AMP-dependent voltage-sensitive ion channels. After chronic opioid exposure of DRG/spinal cord explants the DRG neurons become tolerant to the acute inhibitory (action potential-shortening) effects of high (uM) concentrations of opioids and remarkably sensitive to the excitatory (action potential-prolonging) effects of low (<pM) opioid concentrations. Similar opioid excitatory supersensitivity is elicited by acute exposure of naive DRG neurons to GM1 ganglioside. DRG neurons in culture will be studied by current-clamp and voltage-clamp recordings to test the hypothesis that: (1) opioid excitatory supersensitivity which develops in chronic opioid-treated sensory neurons and renders the cells opioid-dependent is due to enhanced efficacy of Gs-coupled excitatory opioid receptor functions following up-regulation of cAMP-dependent GM1 ganglioside levels; (2) tolerance is due, in part, to phosphorylation of inhibitory opioid receptors by cAMP-dependent protein kinase that uncouples these receptors from Gi/Go during sustained agonist binding. Tests will be made to block the development of dependence in chronic opioid-treated DRG neurons by co-treatment with agents that selectively interfere with elevation of GM1 ganglioside, e.g. cholera toxin-B subunit or glycosyltransferase inhibitors, and to determine if such treatments also attenuate development of tolerance to the inhibitory effects of opioids. Conversely, opioid excitatory supersensitivity in DRG neurons will be selectively induced by chronic exposure to much lower (nM) concentrations of opioids in order to determine if dependence can occur in the absence of tolerance. Correlative biochemical assays of GM1 and cAMP levels, will be made during chronic treatment of DRG neurons with high and low concentrations of specific mu, delta and kappa agonists, as well as binding assays to test for down- or up-regulation of inhibitory or excitatory opioid receptors, respectively. Whole-cell voltage-clamp and single- channel patch-clamp analyses will be made on chronic opioid-treated DRG neurons to determine if specific subtypes of K+ and Ca2+ channels show plastic alterations in functional properties after development of tolerance and/or dependence. These studies may provide new methods for preventing some types of opiate addiction by treatments that block upregulation of GM1 ganglioside in the affected neurons.